Abstract: The evolution of myocardial injury during ischemia reperfusion remains incompletely understood. It is now well documented that a cardiomyocyte (CM) can react to injury by undergoing necrosis, apoptosis or autophagy. However, the full significance of these various forms of cardiomyocyte (CM) injury, as well as their interaction with each other, remains poorly understood. The interplay between cardiomyocyte (CM) apoptosis and autophagy is of particular interest since both are highly regulated and energy requiring processes that are amenable to modulation during reperfusion. The overall aim of this proposal is thus to use novel molecular and microstructural imaging techniques, recently developed in our centers, to image cardiomyocyte apoptosis and autophagy in vivo. By using these techniques we aim to better understand how programmed CM death occurs during myocardial reperfusion and how to prevent this. The central hypothesis of this proposal is that CMs in the midmyocardium are the most susceptible to apoptosis due to the balance of ischemia and reperfusion that exist in this zone, but that they are also the most amenable to salvage through the development of autophagy, which protects the CM by removing dysfunctional mitochondria and other pro-apoptotic stimuli. Using AnxCLIO-Cy5.5 for MRI and fluorescence imaging of CM apoptosis, a near-infrared fluorochrome activated by the lysosomal cathepsins involved in CM autophagy, and diffusion spectrum MRI to visualize myocardial fiber architecture we aim to: 1) determine how the transmural variation in the severity of ischemia and the degree of reperfusion influence the development of cardiomyocyte apoptosis and autophagy, 2) study the effects of postconditioning on cardiomyocyte apoptosis and autophagy 3) determine the impact of cardiomyocyte apoptosis and autophagy on the integrity of myocardial fiber architecture. The proposed imaging approach is highly translational and will allow aspects of CM loss at the cellular level to be correlated with readouts of myocardial structure and function that can be obtained in both mice and humans. The proposed study has the potential to impact clinical care significantly by facilitating the development of strategies to increase the salvage midmyocardial cardiomyocytes, and thus convert highly transmural and poorly tolerated myocardial infarcts into well-tolerated subendocardial infarcts. Narrative: The myocardium (heart muscle) can respond to a lack of blood supply in several ways, some of which may be protective and others deleterious. We aim in this proposal to use novel molecular and microstructural imaging techniques to better understand the response of the myocardium to acute injury, and thus to facilitate the development of novel cardio-protective strategies. The development of such strategies has the potential to significantly reduce the rapidly rising number of patients with heart failure, and is thus of major clinical and public health significance.